Background: HER2 overexpression may be a prognostic factor for poor outcomes in pts with high-risk UC. Publications report a wide variability of HER2 expression in UC, with ≥2+ HER2 expression by immunohistochemistry (IHC) reported in <10% to >50% of cases. DN24-02 is an investigational autologous immunotherapy targeting HER2, based on the same manufacturing platform used for sipuleucel-T. NeuACT (N10-1; NCT01353222) is designed to evaluate whether DN24-02 can prolong survival when given as adjuvant therapy following surgical resection in pts with high risk HER2-expressing UC (Bajorin, et al. ASCO 2012). Here we report preliminary data for HER2 expression on primary tumor and positive lymph node samples, and covariate analyses. Methods: Trial eligibility criteria include surgical resection of a primary UC, with either ≥pT2 or pN+ staging, and HER2 expression ≥1+ IHC. Surgical specimens are screened for HER2 expression by central pathology laboratory review and HER2 positivity is scored using the Dako HercepTest system. Results: As of December 2012, tumor specimens from 114 pts have been screened. Of these pts, 84 (74%; 95% CI: 65–82%) had a HER2 expression score ≥1+ in the primary tumor, with 36 (32%) having a 2+ score and 5 (4%) having a 3+ score. Thirty-eight pts also had HER2 expression levels evaluated in lymph node samples. Of these 38 pts, 35 (92%; 95% CI: 79–98%) had a HER2 expression score ≥1+ in the lymph nodes, with 17 (45%) having a 2+ score and 4 (11%) having a 3+ score. Gender, primary tumor site, nodal stage and prior neoadjuvant chemotherapy were not significantly correlated with HER2 expression (p≥0.10). Conclusions: To date, high frequencies (≥74%) of HER2 expression ≥1+ in primary tumor and lymph node samples of UC pts have been observed. No baseline variables, including prior neoadjuvant chemotherapy, appear to affect the rate of HER2 expression. Although preliminary, these data are consistent with prior studies that have noted a higher incidence of HER2 expression in UC lymph node tumor vs. primary tumor and suggest that HER2 protein expression is common in high-risk UC. Clinical trial information: NCT01353222.